Juvenile hormones (JH) regulate a wide variety of developmental and physiological processes in insects. Comparison of microarray data on JH-induced genes in the fruit fly, Drosophila melanogaster, L57 cells and in the honey bee, Apis mellifera, identified 16 genes that are induced in both species. Analysis of promoter regions of these 16 D. melanogaster genes identified DmJHRE1 (D. melanogaster JH response element 1). In L57 cells, the reporter gene regulated by DmJHRE1 was induced by JH III. Two proteins (FKBP39 and Chd64) that bind to DmJHRE1 were identified. FKBP39 and Chd64 double-stranded RNA inhibited JH III induction of a reporter gene regulated by DmJHRE1. FKBP39 and Chd64 proteins expressed in yeast bound to DmJHRE1. Two-hybrid and pull-down assays showed that these two proteins interact with each other as well as with ecdysone receptor, ultraspiracle, and methoprene-tolerant protein. Developmental expression profiles and JH induction of mRNA for FKBP39 and Chd64 proteins and their interaction with proteins known to be involved in both JH (methoprene-tolerant protein) and ecdysteroid action (ecdysone receptor and ultraspiracle) suggest that these proteins probably play important roles in cross-talk between JH and ecdysteroids.